China Animal Husbandry and Veterinary Medicine ›› 2023, Vol. 50 ›› Issue (3): 1150-1159.doi: 10.16431/j.cnki.1671-7236.2023.03.029

• Preventive Veterinary Medicine • Previous Articles     Next Articles

Isolation,Identification and Genome Sequence Analysis of Bovine Viral Diarrhea Virus LN-1 Strain

LIU Kexin1, WANG Chao2, ZHANG Ao1, LIU Jiali1, TAN Bin1, ZHANG Shuqin1   

  1. 1. Institute of Special Animals and Plants, Chinese Academy of Agricultural Sciences, Changchun 130122, China;
    2. Heilongjiang Academy of Agricultural Sciences, Harbin 150086, China
  • Received:2022-09-22 Online:2023-03-05 Published:2023-03-02

Abstract: 【Objective】 The purpose of this experiment was to identify and master the epidemiological characteristics and biological characteristics of Bovine viral diarrhea virus (BVDV) in Liaoning province,and to provide theoretical basis and technical support for the prevention and treatment of BVDV.【Method】 The tissues suspected of BVDV infection from a cattle farm in Liaoning province were collected,Infectious bovine rhinotracheitis virus (IBRV),Bovine respiratory syncytial virus (BRSV),and Bovine parainfluenza virus type 3 (BPIV3) and BVDV gene-specific primers were identified by RT-PCR,and MDBK cells were inoculated with interstitial fluid to isolate the virus.After cytopathic observation of the cells,the virus was identified again by electron microscopy,indirect immunofluorescence assay (IFA),and the whole genome sequence of the virus was amplified by PCR and analyzed for genetic evolution.【Result】 The identification result of BVDV specific primer RT-PCR was positive.There was a band at 280 bp,which was consistent with the expected band size.The other pathogen specific primers did not amplify the band,which proved that other pathogens were not infected.The isolated strain did not show obvious cytopathic effect in MDBK cells.After virus purification,virus particles with a diameter of about 50 nm were observed by electron microscopy.The results of IFA showed that bright green fluorescence could be observed in the MDBK cells inoculated with the isolated virus strain,while no fluorescence was observed in the control cells.The isolated virus was named LN-1 strain.The size of the whole genome amplification sequence was 12 269 bp.Genetic evolution analysis showed that LN-1 strain was in the same branch as XC,SD-15 and ZM-95 strains,and the nucleotide similarity with XC strain was 96.2%,belonging to BVDV-1m type.【Conclusion】 In this study,one strain of 1m BVDV strain was successfully isolated,which was of great significance for the follow-up vaccine research and development,epidemiological investigation and pathogenesis research.

Key words: Bovine viral diarrhea virus (BVDV); whole genome sequencing; virus isolation; immunofluorescence identification; genetic evolution analysis

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